Tropical Medicine and

Infectious Disease

Article
A 12-Year Retrospective Study of Invasive Amoebiasis
in Western Sydney: Evidence of Local Acquisition
Ana Domazetovska 1,2,3 , Rogan Lee 4,5 ID , Chandra Adhikari 6 , Matthew Watts 2,4,5 ,
Nicole Gilroy 2 , Damien Stark 7 and Shobini Sivagnanam 1, *
1 Department of Infectious Diseases, Blacktown Hospital, Blacktown NSW 2148, Australia;
Ana.Domazetovska@health.nsw.gov.au
2 Department of Infectious Diseases, Westmead Hospital, Westmead NSW 2145, Australia;
Matthew.Watts@health.nsw.gov.au (M.W.); Nicky.Gilroy@health.nsw.gov.au (N.G.)
3 Department of Microbiology, Liverpool Hospital, Liverpool NSW 2170, Australia
4 New South Wales Health Pathology, Institute of Clinical Pathology and Medical Research,
Westmead Hospital, Westmead NSW 2145, Australia; Rogan.Lee@health.nsw.gov.au
5 The University of Sydney Medical School, Westmead Hospital, Westmead NSW 2145, Australia
6 Department of Anatomical Pathology, Institute of Clinical Pathology and Medical Research,
Westmead Hospital, Westmead NSW 2145, Australia; Chandra.Adhikari@health.nsw.gov.au
7 Department of Microbiology, St. Vincent’s Hospital, Darlinghurst NSW 2010, Australia;
damien.stark@svha.org.au
* Correspondence: shobini.sivagnanam@health.nsw.gov.au; Tel.: +61-9881-8000




Received: 13 May 2018; Accepted: 20 June 2018; Published: 26 June 2018


Abstract: In Australia, amoebiasis is thought to occur in travellers, immigrants from endemic areas,
and among men who have sex with men. Prevalence of amoebiasis in communities with immigrants
from Entamoeba histolytica-endemic countries is unknown. The present study is a retrospective case
series analysis of patients with laboratory-confirmed amoebiasis from Western Sydney Local Health
District, Australia, between years 2005 and 2016. Forty-nine patients with amoebiasis were identified,
resulting in an estimated annual incidence of up to 1.1 cases per 100,000 adults. Many were born in
Australia (15/47) and India (12/47). Three patients (3/37) had no history of overseas travel, two others
had not travelled to an endemic country, and an additional two had a very remote history of overseas
travel; one died of fulminant amoebic colitis. Three patients (3/16) were employed in the food industry
and one had a history of colonic irrigation in an Australian ‘wellness clinic’. Patients had invasive
amoebiasis with either liver abscess (41/48) or colitis (7/48), diagnosed most commonly by serology.
Invasive procedures were common, including aspiration of liver abscess (28/41), colonoscopy (11/49),
and partial hepatectomy (1/49). Although rare, local acquisition of amoebiasis occurs in Western
Sydney and contributes to significant morbidity and hospital admissions.

Keywords: amoebiasis; Entamoeba histolytica; endemic; local acquisition; colitis; liver abscess

1. Introduction
Amoebiasis, caused by the parasite Entamoeba histolytica, has a worldwide distribution, with an
estimated 50 million people being infected [1]. With 40,000–100,000 deaths reported yearly, it is the
second leading cause of death from parasitic diseases worldwide [2]. High-risk areas include South
Asia, Southeast Asia, the Middle East, and South America [3]. E. histolytica is acquired through
ingestion of food or water contaminated with faecal cysts; sexual transmission has also been reported,
particularly via contact with commercial sex workers [4] or in men who have sex with men (MSM),
probably via faecal-oral route [5]. Clinical features of amoebiasis include asymptomatic colonisation,

Trop. Med. Infect. Dis. 2018, 3, 73; doi:10.3390/tropicalmed3030073 www.mdpi.com/journal/tropicalmed

Trop. Med. Infect. Dis. 2018, 3, 73 2 of 8

which can progress to invasive disease in the form of amoebic colitis or extra-intestinal disease, with the
most common manifestation being a liver abscess [6].
In Australia, amoebiasis is thought to be imported by travellers and recent immigrants from
high risk areas [7], with an estimated prevalence of 2% in travellers and 1% in immigrants [8].
Locally-acquired disease affects the MSM population [5], and the Indigenous community in northern
Australia [9]. As amoebiasis is not a nationally notifiable disease in Australia, the prevalence of
amoebiasis, including the prevalence of locally-acquired cases, is unknown. In this study, we aim
to determine the epidemiology of amoebiasis in Western Sydney, an area with a high proportion of
immigrants from endemic countries [10], in order to raise awareness and improve the morbidity and
mortality associated with invasive disease.

2. Materials and Methods

2.1. Study Setting

Western Sydney Local Health District (WSLHD) covers four public hospitals that predominantly
see adult patients: Westmead, Blacktown, Auburn, and Mount Druitt hospitals. In 2013, the district
had an estimated population size of 876,500; 43% of the residents were born overseas and 45% spoke a
language other than English at home in 2011 [10].

2.2. Study Cohorts

All patients from WSLHD who had a positive amoebic serology, stool microscopy suggestive of
E. histolytica/dispar, and/or a positive E. histolytica polymerase chain reaction (PCR) between years
2005 and 2016 were identified using the laboratory information system and were eligible for inclusion
in the study. Electronic and paper medical records of eligible patients were retrospectively reviewed to
determine demographics, clinical features, and outcome data. Ethics approval was obtained from the
WSLHD human research ethics committee.

2.3. Diagnostic Methods

2.3.1. Microscopy
Stool microscopy looking specifically for ova, cysts, and parasites (OCP) were performed
upon clinicians’ requests. Stool specimens were received either fresh or in sodium acetate-acetic
acid-formalin (SAF) fixative. When there was no travel history provided, all unstained stool
samples underwent microscopic examination (wet preparation) for leucocytes and erythrocytes and
subsequently a lateral flow assay was performed for giardia and cryptosporidium antigens only.
Apart from a standard wet mount, the laboratory would prepare and examine stool specimens stained
with a modified iron-haematoxylin stain (Para-Stain. Catalogue number MV1284, Oxoid Australia Pty.
Ltd., Thebarton, South Australia) when a relevant travel history was provided and/or when a specific
request for E. histolytica examination was received.

2.3.2. Serology
Patients’ sera were batched and E. histolytica serology was performed once per week at the
Centre for Infectious Diseases and Microbiology Laboratory Services, Westmead, using an indirect
haemagglutination test (IHA) according to the manufacturer’s instructions (Cellognost Amoebiasis,
Siemens Healthcare Diagnostics Products GmbH, Marburg, Germany from 2005 to May 2013; ELI.H.A
Amoeba, ELITech Microbio, Signes, France from May 2013 to November 2015). The result of the IHA
test was reported as a haemagglutination titre. Titres greater than 64 using the Siemens test and greater
than 80 using ELITech were regarded as positive.
Trop. Med. Infect. Dis. 2018, 3, 73 3 of 8

2.3.3. PCR
E. histolytica PCR was performed at St Vincent’s Hospital, Darlinghurst, upon clinician request.
A laboratory-validated in-house PCR targeting the 18S rRNA genes of E. histolytica was used to identify
E. histolytica [11] between 2005 and November 2015. This was subsequently replaced with RT-PCR kit,
Easy Screen Enteric Protozoan Detection Kit (Genetic Signatures, Sydney, Australia) [12].

2.3.4. Histopathology
Tissue samples were referred to the Department of Anatomical Pathology, Institute of Clinical
Pathology and Medical Research at Westmead Hospital. Colonic biopsies were stained with
haematoxylin and eosin; the diagnosis of amoebic colitis was reported when E. histolytica trophozoites
were seen under light microscopy.

2.4. Definitions
Asymptomatic colonisation was defined as the presence of E. histolytica in stool in the absence
of colitis or extra-intestinal manifestations. Amoebic colitis referred to symptomatic patients with
microbiological or histopathological evidence of amoebiasis, without an established alternate cause.
Amoebic liver abscess (ALA) was defined as any patient with microbiological or histopathological
evidence of amoebiasis with an abscess demonstrated on liver imaging. Patients with positive bacterial
cultures of the aspirated liver abscess or blood with borderline positive amoebic serology were excluded
from the study.
Invasive procedure was defined as having a colonoscopy, peripheral insertion of central catheter
(PICC) for intravenous antibiotics, aspiration of ALA, or having a surgical resection.

3. Results
From a total of 173 patients tested by serology, 63 had positive titres and 49 met the inclusion criteria
for this study. All 49 cases were positive by serological testing. In addition, 4 of 13 serology-positive cases
that had faecal concentrate examination performed were positive for E. histolytica/dispar. E. histolytica
was positive on liver abscess fluid from three patients and faecal samples from two patients when tested
by PCR. Interestingly, these five PCR positive cases were not detected by microscopy. Another four
patients were positive for amoebae on histopathology of colonic biopsy. These four patients had already
been identified as positive by microbiological tests (serology: four; stool microscopy: two). Amongst
the excluded cases, 9 had a clear alternate diagnosis, with 8 of these having borderline positive serology.
Another four had a history of previous treatment for amoebiasis. One had no record for review.

3.1. Demographic Data and Risk Factors (Table 1)

There were more male than female patients; none were HIV positive. Where sexual history was
recorded (n = 9), all were heterosexuals. Many patients were born in Australia and India. Among the
Australian-born patients, 3/15 had never travelled overseas. One was a female hospital volunteer
with fulminant amoebic colitis and sepsis. The other two were men with ALA, one of whom had
previous colonic irrigation at a ‘wellness clinic’ and at home. Two additional men born in Australia
with amoebiasis proven on colonoscopy had only travelled to the USA and Japan. One intermittently
lived with a family member who had travelled to Southeast Asia. Another Australian-born male
patient with ALA had last travelled to Fiji 30 years prior. One male patient with ALA was born in
Malta and had no overseas travel for 46 years since migration. Occupation was recorded for 16 patients,
with three cases employed in the food industry (Table 1).

3.2. Clinical Features and Investigations (Table 2)

The three main presenting symptoms were fever, right upper quadrant pain, and diarrhoea.
There were 41 cases of ALA. Seven had amoebic colitis. The eosinophil count was measured in
Trop. Med. Infect. Dis. 2018, 3, 73 4 of 8

48 patients and was elevated in 13, with a range of 0.6–3.2 × 109 /L. All ALA was located within the
right lobe, with a single lesion, except for two patients who had multiple abscesses within the right
lobe. The size of the abscesses varied, with the largest measuring 15 cm in diameter (Table 2).

Table 1. Demographic data of patients with invasive amoebiasis at Western Sydney Local Health
District between 2005 and 2016.

Demographics No. of Cases *

Male 43/49 (88%)
Age (median; interquartile range) (49; 34, 61)
Country of Birth
Australia 15/47 (32%)
India 12/47 (26%)
Other † 19/47 (40%)
Travel History
No overseas travel 3/37 (8%)
Remote travel only 2/37 (5%)
Travel to non-endemic countries only 2/37 (5%)
Employed in food industry 3/16 (19%)
* The denominators refer to the number of patients in whom this information was recorded. † Philippines (n = 1),
Afghanistan (n = 3), England (n = 3), Pakistan (n = 2), Vietnam (n = 1), Poland (n = 1), Serbia (n = 2), Burma (n = 1),
Fiji (n = 2), China (n = 1), Samoa (n = 2), Sri Lanka (n = 1), Malta (n = 1).

Table 2. Clinical features, diagnostic testing, management, and outcome data of patients with invasive
amoebiasis managed at Western Sydney Local Health District between 2005 and 2016.

Clinical Features No. of Cases

Fever 39/49 (80%)
Right upper quadrant pain 35/49 (71%)
Diarrhoea 17/49 (35%)
Positive Diagnostic Investigations
Serology 49/49 (100%)
PCR (liver aspirate) 3/6 (50%)
PCR (Stool) 2/3 (67%)
Stool microscopy 4/13 (31%)
Histopathology 4/11 (36%)
Diagnosis *
Liver abscess 41/48 (85%)
Amoebic colitis 7/48 (15%)
Management and Outcomes
Admission
Surgical 23/45 (51%)
Medical 22/45 (49%)
Not admitted 4/49 (8%)
Treatment
Received an antibiotic active against E. histolytica prior to diagnosis 43/49 (88%)
Received a luminal agent 30/49 (61%)
Invasive procedure 36/49 (73%)
Aspiration of hepatic abscess 28/41 (68%)
Partial hepatectomy 1/49 (2%)
Colonoscopy 11/49 (22%)
Peripherally inserted central cannula 4/49 (8%)
Death 1/49 (2%)
* Type of amoebiasis could not be determined in one patient due to insufficient information. PCR: polymerase
chain reaction.
Trop. Med. Infect. Dis. 2018, 3, 73 5 of 8

3.3. Management
Of the 49 cases with invasive amoebiasis, four patients were seen through an outpatient clinic and
45 were admitted to hospital (Table 2). There were 23 surgical admissions, of which 17 were managed
under the upper gastrointestinal surgical team and six under general surgery. The other 22 cases were
medical admissions, of which 17 were managed under gastroenterology and five under infectious
diseases teams. Most patients (43/49) received antibiotics (metronidazole) active against E. histolytica
before a diagnosis was made, often at a dose lower than that recommended for an ALA. One patient
with no overseas travel history died of fulminant amoebic colitis prior to receiving appropriate therapy;
she was treated empirically with piperacillin/clavulanate and ciprofloxacin instead. This patient
had positive E. histolytica PCR on stool and a high antibody titre (1280) but died prior to the results
being available. Treatment with an intraluminal agent (paromomycin) was documented to be given to
30/49 patients.
Overall, 36/49 patients underwent an invasive procedure, with 28/41 ALA being aspirated.
One patient had partial hepatectomy for the liver lesion, with the diagnosis of an ALA subsequently
made by serology following outpatient referral to an infectious diseases physician. Of the 13 patients
with ALA who did not undergo an invasive procedure, eight were admitted under a medical team,
one was managed as an outpatient, and four were admitted under surgery with infectious diseases
team consulting. These 13 patients had ALA with a range of sizes, including an abscess of 15 cm
diameter with no complications recorded. There was no history of relapsed amoebiasis documented in
any of the 49 patients.

4. Discussion
This present study is the first epidemiological study looking at invasive amoebiasis in New
South Wales, Australia, where E. histolytica is not considered to be endemic. We found that although
amoebiasis was associated with significant morbidity and mortality, it was a rare infection with an
estimated annual incidence of up to 1.1 cases per 100,000 adults in ethnically-diverse Western Sydney
(Figure 1). This was likely an underestimate, as it did not capture cases diagnosed in the private sector.
The majority of our cases had invasive amoebiasis with a liver abscess. Microscopy is an insensitive
test for detection of E. histolytica [13], and PCR testing was referred to an external laboratory upon
clinician request. As a result, our main selection criterion was positive serology using IHA method.
This test is known to have a specificity of over 90% and a high sensitivity for invasive amoebiasis [14],
with the most common manifestation being ALA. In addition, only patients with documented travel
history had stool concentration (Mini Parasep SF Faecal Parasite Concentrator, Apacor, Workingham,
England) and microscopy for E. histolytica; this was likely to have had a negative impact on the rate
of detection of amoebic colitis. We were also unable to establish the rate of carriage of E. histolytica
cysts/trophozoites in our patients with ALA, as stool tests were not done in most cases. Finally, as our
data largely captured symptomatic patients seen in the hospital setting, there may be a greater burden
of asymptomatic colonization in Western Sydney that remains undiagnosed.
With at least three patients with no previous travel history diagnosed with amoebiasis, our study
provides evidence that amoebiasis can be acquired locally in Australia. Previous studies have only
shown evidence for local acquisition in northern Australia [9,15] and within certain population groups
such as MSM [11]. An additional two cases in our study had only travelled to the USA and Japan
and two others had a very remote travel history. Although prolonged latency period could not be
excluded [16], these cases probably reflect more recent local acquisitions. Hence, amoebiasis should be
considered in the differential diagnosis of patients presenting with liver abscesses or colitis, even if
there is no travel history.
Trop. Med. Infect. Dis. 2018, 3, 73 6 of 8
Trop. Med. Infect. Dis. 2018, 3, x FOR PEER REVIEW    6 of 8 

 
Figure  1.  Incidence  of  invasive  amoebiasis  at  Western  Sydney  Local  Health  District  (WSLHD) 
Figure 1. Incidence of invasive amoebiasis at Western Sydney Local Health District (WSLHD) between
between years 2005 and 2016. Population size estimates in WSLHD per year were derived from the 
years 2005 and 2016. Population size estimates in WSLHD per year were derived from the following
following website: http://www.healthstats.nsw.gov.au/. Patients 15 years and over were considered 
website: http://www.healthstats.nsw.gov.au/. Patients 15 years and over were considered adults.
adults. 

Inn our
our population,
population,  there
there  was
was  aa  high
high  incidence
incidence  of of  invasive
invasive  procedures
procedures  among
among  patients with
patients  with 
amoebiasis, including
amoebiasis,  including aspiration
aspiration  ofof 
liver abscesses
liver  andand 
abscesses  one case
one  of partial
case  hepatectomy,
of  partial  despite despite 
hepatectomy,  studies
showing medical management alone leading to complete recovery [17,18].
studies showing medical management alone leading to complete recovery [17,18]. One of the factors  One of the factors that
contributed to diagnostic delays was a lack of consideration of the diagnosis.
that contributed to diagnostic delays was a lack of consideration of the diagnosis. Due to the tests  Due to the tests
being batched, even when the possibility of amoebiasis was raised, there were delays in obtaining
being batched, even when the possibility of amoebiasis was raised, there were delays in obtaining 
results. In some cases, invasive procedures were performed prior to the availability of confirmatory
results. In some cases, invasive procedures were performed prior to the availability of confirmatory 
results. Close 
results.  Close liaison 
liaison with 
with the 
the microbiology 
microbiology laboratory 
laboratory may  may expedite 
expedite testing 
testing and 
and facilitate 
facilitate early 
early
diagnosis in such cases. Given that misdiagnosis and delayed diagnosis
diagnosis in such cases. Given that misdiagnosis and delayed diagnosis can lead to fulminant and  can lead to fulminant and
necrotising colitis [19], and in some cases, death [20], empiric therapy with high-dose
necrotising colitis [19], and in some cases, death [20], empiric therapy with high‐dose metronidazole  metronidazole is
of of 
is  paramount
paramount  importance
importance in all
in patients, irrespective
all  patients,  of travel
irrespective  history,
of  travel  pending
history,  diagnostic
pending  test results.
diagnostic  test 
Amoebiasis
results.  is not a nationally notifiable disease in Australia. However, our study raises a number
of public health concerns.
Amoebiasis  Firstly, three
is  not  a  nationally  patients disease 
notifiable  were employed in theHowever, 
in  Australia.  food industry. Food handlers
our  study  raises  a 
are well represented in the literature as an at-risk group for transmission of
number of public health concerns. Firstly, three patients were employed in the food industry. Food  parasitic infections [21–23].
For this reason, regular stool screening for parasites in food handlers is practiced in
handlers  are  well  represented  in  the  literature  as  an  at‐risk  group  for  transmission  of  parasitic  some endemic
countries [23,24] and demonstration of stool clearance is required in others [25]. Another public
infections [21–23]. For this reason, regular stool screening for parasites in food handlers is practiced 
health concern raised in our study was the risk of amoebiasis transmission during colonic irrigation.
in some endemic countries [23,24] and demonstration of stool clearance is required in others [25]. 
One patient with amoebiasis had never travelled overseas but had undergone colonic irrigation,
Another public health concern raised in our study was the risk of amoebiasis transmission during 
a reported
colonic  source One 
irrigation.  of amoebiasis outbreaks
patient  with  [26].had 
amoebiasis  Public
never health involvement
travelled  overseas would help
but  had  identify
undergone 
and treat potential point source outbreaks from such practices. Finally, one
colonic irrigation, a reported source of amoebiasis outbreaks [26]. Public health involvement would  patient with no known
history
help  of travel
identify  to treat 
and  endemic countries
potential  had
point  regular
source  contact with
outbreaks  from a family member from
such  practices.  Southeast
Finally,  Asia.
one  patient 
Contact tracing may be necessary to reduce the burden of asymptomatic colonisation,
with  no  known  history  of  travel  to  endemic  countries  had  regular  contact  with  a  family  member  including the
spread of infections between household contacts and family members.
from  Southeast  Asia.  Contact  tracing  may  be  necessary  to  reduce  the  burden  of  asymptomatic 
Our study has several limitations. This retrospective study has missing data on important risk
colonisation, including the spread of infections between household contacts and family members. 
factors for local acquisition of amoebiasis such as sexual practices and, in some instances, full travel
Our study has several limitations. This retrospective study has missing data on important risk 
history. We were not able to capture the true prevalence of amoebiasis in Western Sydney, as our data
factors for local acquisition of amoebiasis such as sexual practices and, in some instances, full travel 
was limited to adult patients reviewed at public hospitals only. Given the small numbers, we were not
history. We were not able to capture the true prevalence of amoebiasis in Western Sydney, as our 
able to perform meaningful statistical analyses to identify risk factors that may be associated with local
data was limited to adult patients reviewed at public hospitals only. Given the small numbers, we 
acquisition
were  of amoebiasis.
not  able  to  perform  Despite these limitations,
meaningful  we believe
statistical  analyses  to that our study
identify  risk highlights amoebiasis
factors  that  may  be 
associated with local acquisition of amoebiasis. Despite these limitations, we believe that our study 
highlights  amoebiasis  as  a  neglected  disease  of  the  Western  world  associated  with  significant 
Trop. Med. Infect. Dis. 2018, 3, 73 7 of 8

as a neglected disease of the Western world associated with significant morbidity and mortality and
emphasizes the need to have a high index of suspicion even in patients without a travel history.

5. Conclusions
Our study suggests that amoebiasis is a significant cause of morbidity and hospital admissions in
Western Sydney, perhaps greater than currently recognised and estimated by this study. There is a
need for greater awareness and consideration of this diagnosis so that management can be improved
and invasive procedures avoided. Diagnosis could also be improved by early testing and result
notification in suspected cases. Our findings indicate that amoebiasis acquisition occurs locally in
certain Australian communities and future studies that identify local risk factors for transmission are
warranted. A prospective multi-centre national surveillance study reporting on demographic and
behavioural risk factors, clinical presentation, laboratory diagnosis, and outcomes would better inform
public health control and clinical management of amoebiasis in the Australian context.

Author Contributions: Conceptualization: A.D. and S.S.; methodology: A.D. and S. S.; validation: R.L. and D.S.;
formal analysis: A.D.; investigation: A.D.; data curation: A.D.; original draft preparation: A.D.; review and editing:
A.D., R.L, C.A., M.W., N.G., D.S., and S.S.; visualization: A.D. and S.S.; supervision: S.S.; project administration:
A.D. and S.S.
Funding: This research received no external funding.
Acknowledgments: We thank Glenys Conner, PathNet Application Specialist, NSW Health Pathology for
data retrieval.
Conflicts of Interest: The authors declare no conflict of interest.

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